, Volume 68, Supplement 1, pp 101–105 | Cite as

Measurement of Acetylcholine in Rat Brain Microdialysates by LC–Isotope Dilution Tandem MS

  • Laszlo Prokai
  • Petr Fryčák
  • Stanley M. StevensJr.
  • Vien Nguyen
Full Short Communication


An LC–MS–MS method was developed for measuring acetylcholine (ACh) in an aqueous medium using reversed-phase ion-pair chromatography, electrospray ionization on a quadrupole ion trap instrument and a tetradeuterated analogue (ACh-1,1,2,2-d4) as an internal standard. A rapid separation was achieved on a 5-cm long octadecylsilica column (2.1 mm i.d.) by employing heptafluorobutyric acid (0.1% v/v) as an ion-pairing agent and requiring 10% v/v acetonitrile in 20 mM ammonium formate buffer under isocratic elution at 200 μl min−1 flow rate. The instrument’s response was calibrated with samples containing known mole ratios of ACh and ACh-1,1,2,2-d4 in an artificial cerebrospinal fluid, which afforded the conclusion that analyte concentrations could be determined by multiplying the measured analyte to internal standard ion-current ratio with the known molar concentration of the ACh-1,1,2,2-d4 added. The rapid and simple assay was tested by measuring the basal neurotransmitter concentration in rat brain microdialysates without the use of a cholinesterase inhibitor upon sample collection.


Column liquid chromatography–tandem mass spectrometry Electrospray ionization Isotope dilution In vivo intracranial microdialysis Acetylcholine 


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Copyright information

© Vieweg+Teubner | GWV Fachverlage GmbH 2008

Authors and Affiliations

  • Laszlo Prokai
    • 1
  • Petr Fryčák
    • 1
  • Stanley M. StevensJr.
    • 1
  • Vien Nguyen
    • 1
  1. 1.Department of Molecular Biology and ImmunologyUniversity of North Texas Health Science CenterFort WorthUSA

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